The primary goal of the research proposed in this grant application is to identify and characterize genes that modify susceptibility to skin tumor promotion using the multi-stage skin carcinogenesis model in mice. Much data from both human epidemiologic and animal studies support the hypothesis that cancer susceptibility in the general population is a function of multiple, poorly penetrant modifier genes that control the propensity toward environmental carcinogen-induced tumor development. Data suggest that susceptibility to the tumor promotion stage is a major determinant of overall susceptibility to multi-stage, epidermal carcinogenesis in mice. Identifying and characterizing genes that modify susceptibility to tumor promotion is crucial for a complete understanding of multistage carcinogenesis and for developing effective cancer prevention strategies. Genes that modify susceptibility and severity of other disease syndromes in humans have successfully been identified in animal models using approaches similar to those proposed in this application to identify and characterize promotion susceptibility loci. To identify genes that modify promotion susceptibility, we have used the classic multi-stage skin tumorigenesis model in the mouse, which is an excellent animal model to study epithelial carcinogenesis in humans. Genetic control of susceptibility to skin tumor promotion by the phorbol ester, 12- O-tetradecanoylphorbol-13-acetate (TPA), in crosses between susceptible DBA/2 and resistant C57BL/6 mice is a multigenic trait and we have mapped promotion susceptibility loci to chromosomes (chr) 1 (Psl3), 2 (Psl2), 9 (Psl1), and 19 (Psl4). Analysis of C57BL/6.Psl1dba congenic mouse strains suggests that at least three genes underlie the effects of Psl1 on skin tumor promotion susceptibility. We have designated these loci as Psl1.1, Psl1.2, and Psl1.3. Furthermore, global gene expression analyses using cDNA microarrays revealed that glutathione S-transferase alpha 4 (Gsta4), which maps within Psl1.2, is expressed at 20-fold higher levels in the epidermis of TPA-treated C57BL/6 compared to DBA/2 mice. Gsta4 is a glutathione-S-transferase (GST) and a major substrate for Gsta4 is 4-hydroxy-2(E)-nonenal (4-HNE), a product of lipid peroxidation. Recent preliminary studies indicate that C57BL/6 mice, null for Gsta4, are more sensitive than wild-type mice to the multi-stage skin tumor protocol, using TPA as a promoter. These observations, taken together with published data supporting a role for Gsta4 in response to oxidative stress, suggests that Gsta4 is a good candidate for a gene that underlies the effect of Psl1.2 on TPA promotion susceptibility. In the proposed research, we will test the hypotheses that Gsta4 modifies the response to TPA skin tumor promotion in DBA/2 and C57BL/6 mice by regulating the level of 4-HNE following TPA treatment. We will further characterize the Psl1.2 locus to determine if any other genes mapping within the locus are modifiers of TPA skin tumor promotion susceptibility. In addition, we will identify and characterize genes mapping to Psl1.1 and Psl1.3 that are associated with responsiveness to TPA-induced skin tumor promotion in DBA/2 and C57BL/6 mice. The Specific Aims are: i) To further characterize the role of Gsta4 as a modifier of TPA skin tumor promotion susceptibility; ii) To characterize the mechanism for, and consequences of, strain-specific induction of Gsta4; and iii) To identify and characterize genes mapping within the Psl1.1 and Psl1.3 loci that modify the response to TPA skin tumor promotion. PUBLIC HEALTH RELEVANCE: The primary goal of the research proposed in this grant application is to identify and characterize genes that modify susceptibility to skin tumor promotion using the multi- stage skin carcinogenesis model in mice. Data generated from these studies will lead to identification of genes that regulate susceptibility to cancer in human populations and the development of novel cancer prevention strategies.